Researchers have employed a variety of techniques in an attempt to impair cell adhesion to a surface extant in a living organism or to be introduced into a living organism. Such techniques have significant medical applications because such techniques can be used to improve the biocompatibility of various procedures, transplants and implants. These techniques, however, have presented a variety of problems making their use in vivo, or for in vivo biologic transplants and implants, inappropriate. The technique described herein can be used in a variety of medical applications. In particular, the technique described herein has numerous applications for vascular procedures and biologic transplants. More specifically, the technique described herein, may be used to decrease blood vessel thrombosis and restenosis, reduce graft thrombosis, reduce the complications of vascular surgery and reduce the immunogenicity of transplanted tissues and cellular groups.
With respect to vascular procedures, researchers have sought to use a variety of methods to inhibit platelet deposition on damaged blood vessels because such deposition leads to thrombosis and restenosis. Restenosis primarily results from smooth muscle cell migration and mitosis in the inner layer of the blood vessel, a process known as intimal hyperplasia. When the luminal surface of a blood vessel is damaged, the subendothelial matrix proteins are exposed which triggers platelet deposition that can lead to acute vessel occlusion. Such damage may also trigger intimal hyper plasia through the release of platelet-derived factors. Platelet-released serotonin and thromboxane A.sub.2 are vasoconstrictors currently thought to be important factors in acute vessel thrombotic occlusion. Platelet-derived growth factor ("PDGF") can stimulate smooth muscle cells to become mitoticaly active. Platelet granule release of PDGF, serotonin and leukotrienes, may also mediate migration of smooth muscle cells to the intimal layer of the vessel. Finally , proteases released from platelets, immune cells and cells damaged by mechanical trauma, may facilitate smooth muscle cell migration by breaking down physical boundaries such as the internal elastic lamina.
Such damage to blood vessels occurs during common vascular procedures such as percutaneous transluminal coronary angioplasty ("PTCA"). PTCA is an alternative to coronary artery bypass grafting, reopening blocked coronary arteries in patients with uncomplicated lesions in one or two vessels. PTCA has a high initial success rate although reocclusion caused by thrombosis and vasoconstriction currently occurs immediately after 2-4% of PTCA procedures. Furthermore, approximately 30-40% of arteries successfully opened by PTCA become reoccluded or restenosed, often within three months. PTCA is performed on more than 300,000 patients in the U.S. annually. Accordingly, modification of the PTCA technique by interrupting platelet deposition onto the injured intimal surface following PTCA, may provide substantial benefit in reducing intimal hyperplasia and coronary restenosis.
Blood vessels are similarly damaged during vascular surgery. Acute thrombotic occlusion at vascular anastomoses is a major complication of microvascular graft placement. Platelets respond to agonists and adhere to collagen and other adhesive proteins present at the anastomotic site resulting in platelet activation and further aggregation. Modification of the anastomotic site that results in a temporary non-thrombogenic coating would afford the anastomosis time to heal and eventually reendothelialize. Furthermore, temporary site specific masking of thrombogenic proteins may greatly reduce acute thrombosis and distal tissue ischemia without the use of systemic antiplatelet agents.
Vascular stent placement in conjunction with coronary angioplasty has grown in popularity as a technique to improve long term vessel patency. Such stents work by providing the vessel with support against mechanical recoil. Stent placement does not inhibit platelet deposition and platelet-mediated hyperplasia and reocclusion by intimal hyperplasia remains a common occurrence with stent placement. Furthermore, the advantages offered by a stent can be offset by increased vascular complications stemming from the stent such as damage to the arterial wall, improper stent openings, and bleeding and dissection at the access site. These complications generally result in longer hospital stays. Accordingly, a technique that permitted development of a coating might be used in lieu of stent placement, or as an adjuvant to stent placement, i.e., one that permitted retention of the significant mechanical benefits of the stent while reducing acute thrombosis and hyperplasia, would be a significant benefit.
A variety of techniques have been employed to inhibit platelet deposition, including systemic delivery of pharmaceuticals. Unfortunately, systemic delivery of pharmaceuticals is associated with an increased risk for bleeding complications.
Investigators also have attempted to inhibit platelet deposition using polymer gels. At least theoretically, because these gels degrade by scission, they create an added risk for embolism during the degradation process. Several in the field have attempted to inhibit platelet deposition and have reported greatly reduced thrombosis on damaged arterial sections coated with a polyethylene glycol ("PEG") gel polymerized with photoactivation and initiators. For example, a technique reported by Hubbell et al. (U.S Pat. No. 5,468,505 to Hubbell et al.) uses light activated eosin Y ("EY") and triethanolamine ("TEA") to initiate polymerization of PEG-diacrylate on vessel surfaces to inhibit surface platelet deposition. Unfortunately, these methods often require numerous steps to accomplish the modification.
The prior art also includes descriptions of covalent modification of suspended proteins with functionalized polyethylene glycol. Unfortunately, the parameters for temperature, pH, time and toxic factors/byproducts for such methodologies are inconsistent with their use with biological tissues.
Others in the field are investigating a variety of genetic engineering strategies to preclude vascular restenosis. Unfortunately, the genetic engineering approaches present significant regulatory and technical hurdles. Genetic transformation often requires exposure of the vessel to viral vectors and may be accompanied by poorly controlled cellular transformation. Furthermore, genetic transformation may induce inflammation, be more expensive, and act less quickly to prevent platelet deposition than the technique described herein.
Researchers have also sought a way to decrease temporarily the immunogenicity of transplanted tissues and cells. Current techniques for immunoisolating transplanted cells (e.g. encapsulation) remain problematic. A modification of a transplanted surface that would decrease the immunogenicity of transplanted cells and tissues would reduce the need for immunosuppressive therapy. Similarly, a surface modification technique might also be used to protect cells on microcarriers in blood or plasma contacting bioreactors. An example of such an application involves the bioartificial liver in which researchers currently use immunoisolative membranes or encapsulation to isolate cells. Transplantation researchers have sought a way to induce host tolerance for transplanted grafts. A technique that masks the immunogenic antigens of the transplanted materials until the host can establish tolerance for cell transplants and whole organ transplants would be a useful conjugate to other methodologies designed to induce tolerance in transplant recipients. Furthermore, a technique that reduces acute thrombosis, fibrinolysis, and complements activation in a donor organ would provide significant clinical benefit.
Thus, a need still exists in the art for an economical and efficient method for modifying tissue and cellular surfaces, under conditions tolerable in vivo, such that they exhibit cell adhesion impairing characteristics. In particular, a need still exists in the art for a method for impairing platelet and leukocyte deposition on vessel surfaces, under conditions tolerable in vivo, following vascular surgery or trauma.
The following abbreviations are used by Applicants throughout the application:
ANOVA=analysis of variance PA0 ECGM=endothelial cell growth media with endothelial growth factors PA0 EDTA=ethylene diaminetetra-acetic acid PA0 EY=eosin Y PA0 HCAEC=human coronary arterial endothelial cells PA0 HT=Hepes-Tyrodes mixture PA0 MW=molecular weight PA0 PEG=polyethylene glycol PA0 PTCA=percutaneous transluminal coronary angioplasty PA0 TEA=triethanolamine